Project Summary/Abstract We propose to upgrade the existing DEER/EPR spectrometer at the University of Washington (UW) to achieve higher sensitivity and higher sample throughput. DEER is a structural technique for the quantification of protein conformational landscapes and protein motions on the nanometer scale. Protein motions underlie the molecular processes at the basis of human life and disease. Therefore, DEER is a crucial technique that provides insights that contribute to the knowledge base necessary for drug development. In combination with X-ray crystallography, NMR and cryo- electron microscopy, DEER is part of a complementary set of experimental biostructural tools. It is especially important for the study of membrane proteins. The number of University of Washington (UW) researchers active in the area of biostructural and molecular biomedical sciences has increased substantially in recent years. There is increased demand for DEER measurement time from a variety of NIH-funded researchers at UW. The existing DEER instrument is old and not able to meet the increased demand due to its inherently low sensitivity. The upgrade requested in this project will (a) dramatically enhance sample throughput to provide more DEER data to more investigators, (b) provide increased resolving power with respect to conformational populations and conformational changes in proteins, (c) provide significantly higher sensitivity that will allow the investigation of proteins and protein complexes at lower concentrations, including full-length membrane proteins. This will expand the scope of systems that can be studied. Research projects that will benefit from the new upgraded DEER capabilities include the study of the molecular mechanisms of allosteric regulation in a broad range of ion channels (HCN, KCNH, TRPV1), the development of new molecular probes for protein kinases from the SRC family, the study of ubiquitin ligases from pathogenic bacteria, and the evaluation of de novo designs of proteins for health applications. Overall, the upgrade will significantly expand the scope of biostructural research at UW, which is conducted in a collaborative multidisciplinary environment.